Peripheral Blood Gene Expression Profiling Reveals Molecular Pathways Associated with Cervical Artery Dissection.

Cervical artery dissection (CeAD) is the primary cause of ischemic stroke in young adults. Monogenic heritable connective tissue diseases account for fewer than 5% of cases of CeAD. The remaining sporadic cases have known risk factors. The clinical, radiological, and histological characteristics of systemic vasculopathy and undifferentiated connective tissue dysplasia are present in up to 70% of individuals with sporadic CeAD. Genome-wide association studies identified CeAD-associated genetic variants in the non-coding genomic regions that may impact the gene transcription and RNA processing. However, global gene expression profile analysis has not yet been carried out for CeAD patients. We conducted bulk RNA sequencing and differential gene expression analysis to investigate the expression profile of protein-coding genes in the peripheral blood of 19 CeAD patients and 18 healthy volunteers. This was followed by functional annotation, heatmap clustering, reports on gene-disease associations and protein-protein interactions, as well as gene set enrichment analysis. We found potential correlations between CeAD and the dysregulation of genes linked to nucleolar stress, senescence-associated secretory phenotype, mitochondrial malfunction, and epithelial-mesenchymal plasticity.

Studying of Molecular Regulation of Developmental Processes of Lower Metazoans Exemplified by Cnidaria Using High-Throughput Sequencing.

A unique set of features and characteristics of species of the Cnidaria phylum is the one reason that makes them a model for a various studies. The plasticity of a life cycle and the processes of cell differentiation and development of an integral multicellular organism associated with it are of a specific scientific interest. A new stage of development of molecular genetic methods, including methods for high-throughput genome, transcriptome, and epigenome sequencing, both at the level of the whole organism and at the level of individual cells, makes it possible to obtain a detailed picture of the development of these animals. This review examines some modern approaches and advances in the reconstruction of the processes of ontogenesis of cnidarians by studying the regulatory signal transduction pathways and their interactions.

Epigenetic-genetic chromatin footprinting identifies novel and subject-specific genes active in prefrontal cortex neurons.

Human prefrontal cortex (PFC) is associated with broad individual variabilities in functions linked to personality, social behaviors, and cognitive functions. The phenotype variabilities associated with brain functions can be caused by genetic or epigenetic factors. The interactions between these factors in human subjects is, as of yet, poorly understood. The heterogeneity of cerebral tissue, consisting of neuronal and nonneuronal cells, complicates the comparative analysis of gene activities in brain specimens. To approach the underlying neurogenomic determinants, we performed a deep analysis of open chromatin-associated histone methylation in PFC neurons sorted from multiple human individuals in conjunction with whole-genome and transcriptome sequencing. Integrative analyses produced novel unannotated neuronal genes and revealed individual-specific chromatin "blueprints" of neurons that, in part, relate to genetic background. Surprisingly, we observed gender-dependent epigenetic signals, implying that gender may contribute to the chromatin variabilities in neurons. Finally, we found epigenetic, allele-specific activation of the testis-specific gene nucleoporin 210 like (NUP210L) in brain in some individuals, which we link to a genetic variant occurring in <3% of the human population. Recently, the NUP210L locus has been associated with intelligence and mathematics ability. Our findings highlight the significance of epigenetic-genetic footprinting for exploring neurologic function in a subject-specific manner.-Gusev, F. E., Reshetov, D. A., Mitchell, A. C., Andreeva, T. V., Dincer, A., Grigorenko, A. P., Fedonin, G., Halene, T., Aliseychik, M., Goltsov, A. Y., Solovyev, V., Brizgalov, L., Filippova, E., Weng, Z., Akbarian, S., Rogaev, E. I. Epigenetic-genetic chromatin footprinting identifies novel and subject-specific genes active in prefrontal cortex neurons.

The ctenophore genome and the evolutionary origins of neural systems.

The origins of neural systems remain unresolved. In contrast to other basal metazoans, ctenophores (comb jellies) have both complex nervous and mesoderm-derived muscular systems. These holoplanktonic predators also have sophisticated ciliated locomotion, behaviour and distinct development. Here we present the draft genome of Pleurobrachia bachei, Pacific sea gooseberry, together with ten other ctenophore transcriptomes, and show that they are remarkably distinct from other animal genomes in their content of neurogenic, immune and developmental genes. Our integrative analyses place Ctenophora as the earliest lineage within Metazoa. This hypothesis is supported by comparative analysis of multiple gene families, including the apparent absence of HOX genes, canonical microRNA machinery, and reduced immune complement in ctenophores. Although two distinct nervous systems are well recognized in ctenophores, many bilaterian neuron-specific genes and genes of 'classical' neurotransmitter pathways either are absent or, if present, are not expressed in neurons. Our metabolomic and physiological data are consistent with the hypothesis that ctenophore neural systems, and possibly muscle specification, evolved independently from those in other animals.

Genomic identification in the historical case of the Nicholas II royal family.

Accurate unambiguous identification of ancient or historical specimens can potentially be achieved by DNA analysis. The controversy surrounding the fate of the last Russian Emperor, Nicholas II, and his family has persisted, in part, because the bodies of 2 children, Prince Alexei and 1 of his sisters, have not been found. A grave discovered in 1991 contained remains putatively identified as those of the Russian Royal family. However, not all family members were represented. Here, we report the results of genomic analyses of new specimens, the human remains of 2 burned skeletons exhumed from a grave discovered in July 2007, and the results of a comprehensive genomic analysis of remains from the 1991 discovery. Additionally, approximately 117 years old archival blood specimens from Nicholas II were obtained and genotyped, which provided critical material for the specific determination of individual identities and kinship identifications. Results of genotypic analyses of damaged historical specimens were evaluated alongside samples from descendants of both paternal and maternal lineages of the European Royal families, and the results conclusively demonstrate that the recently found remains belong to children of Nicholas II: Prince Alexei and his sister. The results of our studies provide unequivocal evidence that the remains of Nicholas II and his entire family, including all 5 children, have been identified. We demonstrate that convergent analysis of complete mitochondrial genome sequences combined with nuclear DNA profiles is an efficient and conclusive method for individual and kinship identification of specimens obtained from old historic relics.

Neurodevelopmental Syndrome with Intellectual Disability, Speech Impairment, and Quadrupedia Is Associated with Glutamate Receptor Delta 2 Gene Defect.

Bipedalism, speech, and intellect are the most prominent traits that emerged in the evolution of Homo sapiens. Here, we describe a novel genetic cause of an "involution" phenotype in four patients, who are characterized by quadrupedal locomotion, intellectual impairment, the absence of speech, small stature, and hirsutism, observed in a consanguineous Brazilian family. Using whole-genome sequencing analysis and homozygous genetic mapping, we identified genes bearing homozygous genetic variants and found a homozygous 36.2 kb deletion in the gene of glutamate receptor delta 2 (GRID2) in the patients, resulting in the lack of a coding region from the fifth to the seventh exons. The GRID2 gene is highly expressed in the cerebellum cortex from prenatal development to adulthood, specifically in Purkinje neurons. Deletion in this gene leads to the loss of the alpha chain in the extracellular amino-terminal protein domain (ATD), essential in protein folding and transport from the endoplasmic reticulum (ER) to the cell surface. Then, we studied the evolutionary trajectories of the GRID2 gene. There was no sign of strong selection of the highly conservative GRID2 gene in ancient hominids (Neanderthals and Denisovans) or modern humans; however, according to in silico tests using the Mfold tool, the GRID2 gene possibly gained human-specific mutations that increased the stability of GRID2 mRNA.

Complete mitochondrial genome and phylogeny of Pleistocene mammoth Mammuthus primigenius.

Phylogenetic relationships between the extinct woolly mammoth (Mammuthus primigenius), and the Asian (Elephas maximus) and African savanna (Loxodonta africana) elephants remain unresolved. Here, we report the sequence of the complete mitochondrial genome (16,842 base pairs) of a woolly mammoth extracted from permafrost-preserved remains from the Pleistocene epoch--the oldest mitochondrial genome sequence determined to date. We demonstrate that well-preserved mitochondrial genome fragments, as long as approximately 1,600-1700 base pairs, can be retrieved from pre-Holocene remains of an extinct species. Phylogenetic reconstruction of the Elephantinae clade suggests that M. primigenius and E. maximus are sister species that diverged soon after their common ancestor split from the L. africana lineage. Low nucleotide diversity found between independently determined mitochondrial genomic sequences of woolly mammoths separated geographically and in time suggests that north-eastern Siberia was occupied by a relatively homogeneous population of M. primigenius throughout the late Pleistocene.

Chromatin profiling of cortical neurons identifies individual epigenetic signatures in schizophrenia.

Both heritability and environment contribute to risk for schizophrenia. However, the molecular mechanisms of interactions between genetic and non-genetic factors remain unclear. Epigenetic regulation of neuronal genome may be a presumable mechanism in pathogenesis of schizophrenia. Here, we performed analysis of open chromatin landscape of gene promoters in prefrontal cortical (PFC) neurons from schizophrenic patients. We cataloged cell-type-based epigenetic signals of transcriptional start sites (TSS) marked by histone H3-K4 trimethylation (H3K4me3) across the genome in PFC from multiple schizophrenia subjects and age-matched control individuals. One of the top-ranked chromatin alterations was found in the major histocompatibility (MHC) locus on chromosome 6 highlighting the overlap between genetic and epigenetic risk factors in schizophrenia. The chromosome conformation capture (3C) analysis in human brain cells revealed the architecture of multipoint chromatin interactions between the schizophrenia-associated genetic and epigenetic polymorphic sites and distantly located HLA-DRB5 and BTNL2 genes. In addition, schizophrenia-specific chromatin modifications in neurons were particularly prominent for non-coding RNA genes, including an uncharacterized LINC01115 gene and recently identified BNRNA_052780. Notably, protein-coding genes with altered epigenetic state in schizophrenia are enriched for oxidative stress and cell motility pathways. Our results imply the rare individual epigenetic alterations in brain neurons are involved in the pathogenesis of schizophrenia.

Mutational re-modeling of di-aspartyl intramembrane proteases: uncoupling physiologically-relevant activities from those associated with Alzheimer's disease.

The intramembrane proteolytic activities of presenilins (PSEN1/PS1 and PSEN2/PS2) underlie production of ß-amyloid, the key process in Alzheimer's disease (AD). Dysregulation of presenilin-mediated signaling is linked to cancers. Inhibition of the γ-cleavage activities of PSENs that produce Aß, but not the ε-like cleavage activity that release physiologically essential transcription activators, is a potential approach for the development of rational therapies for AD. In order to identify whether different activities of PSEN1 can be dissociated, we designed multiple mutations in the evolutionary conserved sites of PSEN1. We tested them in vitro and in vivo assays and compared their activities with mutant isoforms of presenilin-related intramembrane di-aspartyl protease (IMPAS1 (IMP1)/signal peptide peptidase (SPP)). PSEN1 auto-cleavage was more resistant to the mutation remodeling than the ε-like proteolysis. PSEN1 with a G382A or a P433A mutation in evolutionary invariant sites retains functionally important APP ε- and Notch S3- cleavage activities, but G382A inhibits APP γ-cleavage and Aß production and a P433A elevates Aß. The G382A variant cannot restore the normal cellular ER Ca2+ leak in PSEN1/PSEN2 double knockout cells, but efficiently rescues the loss-of-function (Egl) phenotype of presenilin in C. elegans. We found that, unlike in PSEN1 knockout cells, endoplasmic reticulum (ER) Ca2+ leak is not changed in the absence of IMP1/SPP. IMP1/SPP with the analogous mutations retained efficiency in cleavage of transmembrane substrates and rescued the lethality of Ce-imp-2 knockouts. In summary, our data show that mutations near the active catalytic sites of intramembrane di-aspartyl proteases have different consequences on proteolytic and signaling functions.

Whole-genome sequencing identifies a novel ABCB7 gene mutation for X-linked congenital cerebellar ataxia in a large family of Mongolian ancestry.

X-linked congenital cerebellar ataxia is a heterogeneous nonprogressive neurodevelopmental disorder with onset in early childhood. We searched for a genetic cause of this condition, previously reported in a Buryat pedigree of Mongolian ancestry from southeastern Russia. Using whole-genome sequencing on Illumina HiSeq 2000 platform, we found a missense mutation in the ABCB7 (ABC-binding cassette transporter B7) gene, encoding a mitochondrial transporter, involved in heme synthesis and previously associated with sideroblastic anemia and ataxia. The mutation resulting in a substitution of a highly conserved glycine to serine in position 682 is apparently a major causative factor of the cerebellar hypoplasia/atrophy found in affected individuals of a Buryat family who had no evidence of sideroblastic anemia. Moreover, in these affected men we also found the genetic defects in two other genes closely linked to ABCB7 on chromosome X: a deletion of a genomic region harboring the second exon of copper-transporter gene (ATP7A) and a complete deletion of PGAM4 (phosphoglycerate mutase family member 4) retrogene located in the intronic region of the ATP7A gene. Despite the deletion, eliminating the first of six metal-binding domains in ATP7A, no signs for Menkes disease or occipital horn syndrome associated with ATP7A mutations were found in male carriers. The role of the PGAM4 gene has been previously implicated in human reproduction, but our data indicate that its complete loss does not disrupt male fertility. Our finding links cerebellar pathology to the genetic defect in ABCB7 and ATP7A structural variant inherited as X-linked trait, and further reveals the genetic heterogeneity of X-linked cerebellar disorders.

Studying micro RNA Function and Dysfunction in Alzheimer's Disease.

Alzheimer's disease (AD) is a tragic, progressive, age-related neurological dysfunction, representing one of the most prevalent neurodegenerative disorders in industrialized societies. Globally, 5 million new cases of AD are diagnosed annually, with one new AD case being reported every 7 s. Most recently there has been a surge in the study of the regulatory mechanisms of the AD process, and the particular significance of small non-coding ∼22 ribonucleotide RNAs called micro RNAs (miRNAs). Abundant data have profiled miRNA patterns in healthy, aging brain, in mild cognitive impairment (MCI), and in the moderate- and late-stages of AD. The major mode of action of miRNA is to interact, via base-pair complementarity, with ribonucleotides located within the 3' untranslated region (3'-UTR) of multiple target messenger RNAs (mRNAs), and in doing so decrease the capability of that specific mRNA to be expressed. Many miRNAs are highly cell- and tissue-specific. The human brain appears to use only a highly specific fraction of all known human miRNAs, whose speciation and complexity are defined as a discrete subset of all known small non-coding RNAs (sncRNAs) in the brain. In general, in contrast to normally, aging human brain, in AD a family of pathogenically up-regulated miRNAs appear to be down-regulating the expression certain brain-essential mRNA targets, including key regulatory genes involved interactively in neuroinflammation, synaptogenesis, neurotrophic functions, and amyloidogenesis. These up-regulated, NF-kB-sensitive miRNAs, involved in the innate immune and inflammatory response and synaptic, neurotrophic, and amyloidogenic functions include miRNA-9, miRNA-125b, miRNA-146a, and miRNA-155. Other miRNAs of the miRNA-15/107 family, miRNA-153 and miRNA-190, and others, will be discussed. Overall, this manuscript will review the known contribution of miRNAs to aging brain function and the role they appear to play in the incidence and progression of AD.

Genotype analysis identifies the cause of the "royal disease".

The "royal disease," a blood disorder transmitted from Queen Victoria to European royal families, is a striking example of X-linked recessive inheritance. Although the disease is widely recognized to be a form of the blood clotting disorder hemophilia, its molecular basis has never been identified, and the royal disease is now likely extinct. We identified the likely disease-causing mutation by applying genomic methodologies (multiplex target amplification and massively parallel sequencing) to historical specimens from the Romanov branch of the royal family. The mutation occurs in F9, a gene on the X chromosome that encodes blood coagulation factor IX, and is predicted to alter RNA splicing and to lead to production of a truncated form of factor IX. Thus, the royal disease is the severe form of hemophilia, also known as hemophilia B or Christmas disease.

Conversion and compensatory evolution of the gamma-crystallin genes and identification of a cataractogenic mutation that reverses the sequence of the human CRYGD gene to an ancestral state.

We identified a mutation in the CRYGD gene (P23S) of the gamma-crystallin gene cluster that is associated with a polymorphic congenital cataract that occurs with frequency of approximately 0.3% in a human population. To gain insight into the molecular mechanism of the pathogenesis of gamma-crystallin isoforms, we undertook an evolutionary analysis of the available mammalian and newly obtained primate sequences of the gamma-crystallin genes. The cataract-associated serine at site 23 corresponds to the ancestral state, since it was found in CRYGD of a lower primate and all the surveyed nonprimate mammals. Crystallin proteins include two structurally similar domains, and substitutions in mammalian CRYGD protein at site 23 of the first domain were always associated with substitutions in the structurally reciprocal sites 109 and 136 of the second domain. These data suggest that the cataractogenic effect of serine at site 23 in the N-terminal domain of CRYGD may be compensated indirectly by amino acid changes in a distal domain. We also found that gene conversion was a factor in the evolution of the gamma-crystallin gene cluster throughout different mammalian clades. The high rate of gene conversion observed between the functional CRYGD gene and two primate gamma-crystallin pseudogenes (CRYGEP1 and CRYGFP1) coupled with a surprising finding of apparent negative selection in primate pseudogenes suggest a deleterious impact of recently derived pseudogenes involved in gene conversion in the gamma-crystallin gene cluster.

Human hair growth deficiency is linked to a genetic defect in the phospholipase gene LIPH.

The molecular mechanisms controlling human hair growth and scalp hair loss are poorly understood. By screening about 350,000 individuals in two populations from the Volga-Ural region of Russia, we identified a gene mutation in families who show an inherited form of hair loss and a hair growth defect. Affected individuals were homozygous for a deletion in the LIPH gene on chromosome 3q27, caused by short interspersed nuclear element-retrotransposon-mediated recombination. The LIPH gene is expressed in hair follicles and encodes a phospholipase called lipase H (alternatively known as membrane-associated phosphatidic acid-selective phospholipase A1alpha), an enzyme that regulates the production of bioactive lipids. These results suggest that lipase H participates in hair growth and development.

The Caenorhabditis elegans IMPAS gene, imp-2, is essential for development and is functionally distinct from related presenilins.

Presenilins (PSs) are required for Notch signaling in the development of vertebrates and invertebrates. Mutations in human PS1 and PS2 homologs are a cause of familial Alzheimer's disease (AD). The function of the recently identified ancient family of IMPAS proteins (IMP/SPP/PSH) homologous to PSs is not yet known. We show here that, unlike PSs, IMPs (orthologous C. elegans Ce-imp-2 and human hIMP1/SPP) do not promote Notch (C. elegans lin-12,glp-1) proteolysis or signaling. The knock-down of Ce-imp-2 leads to embryonic death and an abnormal molting phenotype in Caenorhabditis elegans. The molting defect induced by Ce-imp-2 deficiency was mimicked by depleting cholesterol or disrupting Ce-lrp-1 and suppressed, in part, by expression of the Ce-lrp-1 derivate. C. elegans lrp-1 is a homolog of mammalian megalin, lipoprotein receptor-related protein (LRP) receptors essential for cholesterol and lipoprotein endocytosis and signaling. These data suggest that IMPs are functionally distinct from related PSs and implicate IMPs as critical regulators of development that may potentially interact with the lipid-lipoprotein receptor-mediated pathway.